Phleomycin resistance as a dominant selectable marker for plant cell transformation

Tobacco cells are sensitive to bleomycin and phleomycin. The Tn5 and the Streptoalloteichus hindustanus (Sh) bleomycin resistance (Ble) genes conferring resistance to these antibiotics have each been inserted into two plant expression vectors. They are flanked by the nopaline synthase (nos) or the cauliflower mosaic virus (CaMV) 35S promoters on one side, and by the nos polyadenylation signal on the other. These four chimaeric genes were introduced into the binary transformation vector pGA 492, which were thereafter mobilized into Agrobacterium tumefaciens strain LBA 4404. The resulting strains were used to transform Nicotiana tabacum cv. Xanthi nc using the leaf disc transformation procedure. In all cases, phleomycin- and bleomycin-resistant tobacco plants were regenerated from transformed cells under selective conditions; however the highest frequency of rooted plants was obtained when transformation was carried out with the Sh Ble gene under the control of the 35S promoter. Phleomycin resistance was stably transmitted to sexual offspring as a dominant nuclear trait as confirmed by Southern blotting.     

Production of alkaline phosphatase by immobilized growing cells of Escherichia coli excretory mutants

Summary In continuous cultures, alkaline phosphatase was synthesised and excreted for more than 250 h by immobilized growing cells in contrast to free cells for which the excretion decreased after 150 h of culture. This observed increase in alkaline phosphatase synthesis and excretion by immobilized cells may have resulted from growing conditions within the gel beads.Offprint requests to: C. Manin     

Continuous production of gibberellic acid in a fixed-bed reactor by immobilized mycelia of Gibberella fujikuroi in calcium alginate beads

Summary The continuous production of gibberellic acid with immobilized mycelia of Gibberella fujikuroi was maintained over a hundred days in a tubular fixed-bed reactor. Free mycelium at the beginning of the storage phase was harvested from G. fujikuroi shake-flask culture and was immobilized by ionotropic gelation in calcium alginate beads.The continuous recycle production system consisted of a fixed-bed reactor, a container in which the culture medium was heated, stirred and aerated, and valves for sample withdrawal or reactant addition during the first 1320 h (55 days). A two-phase continuous extractor was then added for the last 960 hours (40 days). Free and immobilized mycelium shake-flask cultures with the same strain used in the continuous culture system were also realized to compare growth, maintenance and production parameters. The results show about the same gibberellic acid productivity in both free and immobilized mycelium shakeflask cultures: 0.384 and 0.408 mgGA₃·gBiomass^-1 ·day^-1, respectively, whereas in the continuous system the gibberellic acid production is about twice as large for a similar biomass: 0.768 mgGA₃·gBiomass^-1·day^-1. Several factors affecting the overall productivity of the immobilized systems were found to be: the quality and the quantity of mycelia in the biocatalyst beads and the immobilization conditions.     

A minority of 46,XX true hermaphrodites are positive for the Y-DNA sequence including SRY

Summary A total of 30 cases of 46,XX true hermaphroditism was analysed for Y-DNA sequences including the recently cloned gene for male testis-determination SRY. In 3 cases, a portion of the Y chromosome including SRY was present and, in 2 cases, was localised, to Xp22 by in situ hybridisation. Since previous studies have shown that the majority of XX males are generated by an X-Y chromosomal interchange, the Xp22 position of the Yp material suggests that certain cases of hermaphroditism can arise by the same meiotic event. The phenotype in the 3 SRY-positive cases may be caused by X-inactivation resulting in somatic mosaicism of testis-determining factor expression giving rise to both testicular and ovarian tissues. Autosomal or X-linked mutation(s) elsewhere in the sex-determining pathway may explain the phenotype observed in the remaining 27 SRY-negative cases.     

Genetic determinants of glutamine synthetase inDrosophila melanogaster: A gene for glutamine synthetase I resides in the 21B3-6 region

Recombinational and deletion mapping of electrophoretic variants of the glutamine synthetase I isozyme (GSI) in Drosophila melanogaster locates the gene in the 21B region on the second chromosome. We have conducted a genetic analysis of the region extending cytologically from 21A to 21B4-6. Recessive lethal mutations were generated by ethyl methanesulfonate (EMS) and ethyl nitrosourea (ENU) mutagenesis and by hybrid dysgenesis (HD). These lethals fall into seven functional groups, which were partially ordered by complementation with cytologically defined deficiencies of this region generated by hybrid dysgenesis. Two of the EMS- and two of the ENU-induced lethals fulfill biochemical criteria expected for null alleles of the GSI gene.     

Stereoscopic back-scattered electron imaging of silver-stained proteins in nucleoli

By using simultaneously the AgNOR silver staining method, back-scattered electron imaging mode and stereo-tilt in scanning electron microscopy (SEM), it is possible to observe the nucleus through the cell surface, the nucleolus, and the tri-dimensional distribution of the AgNOR-associated acidic proteins. In C3H10T1:2 cells and their 7-12-dimethylbenz--anthracene-treated transformants, the staining demonstrates several intranucleolar silver-staining granules (SSG), surrounded by a weakly staining region. The SSG may represent the fibrillar center (FC) and the weakly staining region, the fibrillar dense component (FD). This component can link several SSG together to form a rope-like structure. In cells with no visible nucleolus and inactive nucleolar organizer regions (NORs) the silver-staining granules are less numerous, close together and the presumed fibrillar dense components are not visible. The SSG are located more peripheraly, and the weakly staining region and the rope-like structure are less prominent in control cell nucleoli than in transformed cells with a comparatively high rate of RNA synthesis.     

Isoproterenol induces a ribosomal modification in the heart and the skeletal muscle of hamsters

The protein synthesis activity of heart, skeletal muscle and liver polysomes from isoprotenerol-treated and control hamsters has been compared in an in vitro non-inititating translation system. Heart and skeletal muscle polysomes from treated hamsters were less active than controls and required a higher magnesium concentration for optimal protein synthesis. These results suggest that there is a conformational modification in heart and skeletal muscle ribosomes from isoprotenerol-treated hamsters. No such change was observed with ribosomes from the liver of isoproterenol-treated hamsters.